Magnitude Improvement of Molecular Signaling Imaging (RMI)

描述（由申请人提供）：最近对细胞信号通路的定义为多细胞生物协调对环境变化的转录反应提供了新的见解。成像细胞信号在哺乳动物将提供洞察的因素，调节这些信号在体内。电子顺磁共振（EPR）成像提供了新的、高分辨率的小鼠氧图像。分子成像与光学检测具有有限的深度灵敏度，使测量更大的动物困难。其他技术受到各种限制。我们在此提议探索低频EPR成像，具有组织深处的灵敏度，以检测作为细胞信号传导的一个例子，诱导缺氧肽信号分子。EPR图像的独特特征包括低信号背景，小型哺乳动物的高空间分辨率（1毫米或更小），缩放到更大动物的前景以及动物组织深处的灵敏度，使这项技术脱颖而出，并鼓励其探索。缺氧诱导因子（Hypoxia Inducible Factor， HIF）蛋白在调控和协调细胞和机体对缺氧的反应中起着重要作用。这项工作将包括构建EPR自旋探针，将其注射到动物体内，在遇到蛋白酶（MMP2）之前，动物是沉默的。它还将涉及在hif1α结合蛋白启动子上游的MMP2基因构建到病毒载体中的工程。然后，病毒载体将被用于1)感染肿瘤细胞或2)周围正常组织。肿瘤或组织缺氧会刺激hif -1 α结合蛋白的产生，hif -1 α结合蛋白会刺激MMP2的产生，MMP2会激活沉默自旋探针。然后获得EPR氧图像，以关联缺氧水平，缺氧细胞信号在活体动物肿瘤中被触发。这将提供体内缺氧信号阈值的测量。这种技术保证了分辨率和灵敏度的数量级提高

DESCRIPTION (provided by applicant): The recent definition of cellular signaling pathways has provided new insight into the means by which multi-cellular organisms coordinate a transcriptional response to environmental change. Imaging cellular signaling in mammals will provide insight into the factors that modulate these signals in vivo. Electron Paramagnetic Resonance (EPR) imaging has provided novel, high resolution oxygen images in mice. Molecular imaging with optical detection has limited depth sensitivity making measurements in larger animals difficult. Other techniques suffer from various limitations. We propose here to explore low frequency EPR imaging, with sensitivity deep in tissues, to detect, as an example of cellular signaling, the induction of hypoxia peptide signal molecules. Unique features of EPR images including low signal background, high spatial resolution (1 mm or less) in small mammals, the promise of scaling to larger animals and sensitivity deep in animal tissues distinguish this technique and encourage its exploration. Hypoxia Inducible Factor (HIF) proteins play a major role in controlling and coordinating the response of cells and organisms to hypoxia. The work will involve the construction of an EPR spin probe, to be injected into the animal, which is silent until it encounters a proteinase (MMP2). It will also involve the engineering of an MMP2 gene construct just upstream of the HIF1alpha binding protein promoter into a viral vector. The viral vector will then be used to 1) infect cells in tumors or 2) surrounding normal tissues. Tumor or tissue hypoxia will stimulate production of HIF-1alpha binding protein which will stimulate the production of MMP2 which will activate the silent spin probe. EPR oxygen images will then be obtained to correlate the levels of hypoxia at which hypoxic cell signaling is triggered in the tumors of a living animal. This will provide a measure of the in vivo threshold for hypoxic signaling. Such a technology promises order of magnitude improvements of resolution and sensitivity